MARCKS silencing differentially affects human vascular smooth muscle and endothelial cell phenotypes to inhibit neointimal hyperplasia in saphenous vein.
Intimal hyperplasia (IH) limits the patency of all cardiovascular vein bypass grafts. We previously found the myristoylated alanine-rich C kinase substrate (MARCKS), a key protein kinase C (PKC) substrate, to be up-regulated in canine models of IH. Here, we further characterize the role of MARCKS in IH and examine the phenotypic consequences of MARCKS silencing by small interfering RNA (siRNA) transfection in human vascular smooth muscle cells (VSMCs) and endothelial cells (ECs) in vitro and use a rapid 10-min nonviral siRNA transfection technique to determine the effects of MARCKS silencing in human saphenous vein cultured ex vivo. We demonstrate MARCKS silencing attenuates VSMC migration and arrests VSMC proliferation in part through the up-regulation of the cyclin-dependent kinase inhibitor p27(kip1). Conversely, MARCKS silencing had little or no effect on EC migration or proliferation. These phenotypic changes culminated in reduced neointimal formation in cultured human saphenous vein. These data identify MARCKS as a pathogenic contributor to IH and indicate therapeutic MARCKS silencing could selectively suppress the "atherogenic," proliferative phenotype of VSMCs without collateral harm to the endothelium. This approach could be readily translated to the clinic to silence MARCKS in vein bypass grafts prior to implantation.
Monahan
Thomas S
TS
Department of Surgery, Division of Vascular Surgery, Beth Israel Deaconess Medical Center, Boston, MA 02115, USA.
Andersen
Nicholas D
ND
Martin
Michelle C
MC
Malek
Junaid Y
JY
Shrikhande
Gautam V
GV
Pradhan
Leena
L
Ferran
Christiane
C
LoGerfo
Frank W
FW
D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D006965:Hyperplasia / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism* / Q000473:pathology*; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D010641:Phenotype; D034741:RNA, Small Interfering / Q000235:genetics*; D012501:Saphenous Vein / Q000378:metabolism* / Q000473:pathology*; D013997:Time Factors; D015854:Up-Regulation
D047908:Intracellular Signaling Peptides and Proteins; C000619710:MARCKS protein, human; D008565:Membrane Proteins; D034741:RNA, Small Interfering; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D050760:Cyclin-Dependent Kinase Inhibitor p27